局部亚低温对脑缺血再灌注治疗时间的影响

Effect of local mild hypothermia on the time of reperfusion of blood

目的观察有效再灌注时间窗内实施局部亚低温对不同时间点血管再通(再灌注)的影响,并探讨其机制。方法将150只雄性大鼠随机分为大脑中动脉闭塞(middle cerebral artery occlusion,MCAO)24h组10只;常温再灌注组50只;MCAO 2h进行亚低温再灌注组50只;MCAO 3h进行亚低温再灌注组40只。动物均于再灌注24h后处死,但MCAO 24h组大脑中动脉闭塞24h后直接处死。采用TTC染色观察梗死体积,用干-湿质量法测定脑含水量,用原位末端标记(TUNEL)观察神经细胞凋亡的变化,采用免疫组化法检测大鼠模型的Bcl-2、Bax及水通道蛋白4(aquaporin4,AQP4)的表达。结果 MCAO 24h组的梗死体积、脑含水量、TUNEL阳性细胞数、及Bcl-2、Bax、AQP4的表达相比较,常温组2～3h有统计学差异(P<0.01),4～6h没有统计学差异(P>0.05);MCAO 2h亚低温再灌注组2～5h有统计学差异(P<0.01),6h没有统计学差异(P>0.05);MCAO 3h亚低温再灌注组3～4h有统计学差异(P<0.01),5～6h没有统计学差异(P>0.05)。亚低温再灌注组各时相点的梗死体积、脑含水量、TUNEL阳性细胞数、Bax、AQP4的表达均显著低于相应常温组,Bcl-2的表达显著高于常温组(P<0.01,P<0.05)。结论实施局部亚低温可延长再灌注治疗时间窗,而且亚低温开始时间越早,其延长时间窗的效果就越显著。这为解决在时间窗内就诊的患者因检查等错过再灌注治疗这一临床难题的解决提供了重要的实验室依据。其机制可能与抑制半暗带细胞的凋亡和改善微循环有关。

Objective To observe the effect of local mild hypothermia applied during window of reperfusion therapy on the different time of reperfusion and its mechanisms.Methods The adult 150 rats were randomly divided into the middle cerebral artery thoroughly obstructed for 24h（MCAO 24h） group（10 rats）,normal temperature groups（NT,50 rats）,MCAO 2h groups（50 rats） and MCAO 3h groups（40 rats）,all rats reperfused for 24h.The expressions of Bcl-2,Bax,aquaporin4（AQP4） were detected by immunohistochemistry.The neuronal apoptosis were dynamically detected by TUNEL.The infarct volumes were evaluated by TTC staining.The water content of rats brain tissue were measured by using dry-wet weight method.Result Compared with MCAO 24h group,in term of the infarction volume and water content of rats brain and the number of apoptotic cells and the expressions of Bcl-2,Bax and AQP4 of normal temperature groups（NT） with 2~3h reperfusion groups of NT had significant difference（P0.01）,groups with 4~6h reperfusion of NT had not significant difference（P0.05）.MCAO 2h applied mild hypothermia group 2~5h reperfusion groups have significant difference（P0.01）,MCAO 6h has not significant difference（P0.05）.MCAO 3h applied mild hypothermia with 3~4h reperfusion groups had significant difference（P0.01）,while MCAO 5~6h had not significant difference（P0.05）.All the groups applied mild hypothermia were lower significantly than NT in term of the infarction volume and water content of rats brain and the number of apoptotic cells and the expressions of Bax and AQP4.MH were higher significantly than NT in term of Bcl-2（P0.01,P0.05）.Conclusion The local mild hypothermia applied during window of reperfusion therapy can prolong the reperfusion therapy.The sooner local mild hypothermia is,the longer the window of reperfusion therapy is.It will provide experimental basis for the clinical problem that the patient visited during the effective window of reperfusion missed the reperfusion therapy due to examination.The reason for that may be that it can inhibit apoptosis,and can better microcirculation.